乏氧激活HIF-1α/miR-210在介入治疗肝癌血管正常化中的机制研究

The tumor angiogenesis is an important prognosis factor in interventional treatment of hepatocelluar carcinoma.HIF-1α/miR-210 pathway may participate in the regulation of angiogenesis in hepatocellular carcinoma. Our previous study demonstrated that tumor vascular normalization existed after treatment of using angiogenesis inhibitor in hepatocellular carcinoma. Based on our preliminary studies, for the down regulation of proangiogenic factor, angiogenesis related factors, and up regulation of angiogenesis inhibitor, we proposed a scientific hypothesis on "The HIF-1α/miR-210 may participate in the regulation of signal pathway in tumor vascular normalization by down regulation of proangiogenic factor, and/or up regulation of angiogenesis inhibitor." In this project, hypoxic hepatoma cell model and hepatoma xenograft nude mice model, morris hepatoma 3924A ACI rat model will be adopted, and the characteristic tumor vascular morphology observed by laser scanning confocal microscope and scanning electron microscope. The HIF-1α/miR-210 pathway will be intervened by siRNA, HIF-1α inhibitor method to study the characteristic and mechanisms of HIF-1α/miR-210 pathway in the regulation of hepatoma vascular normalization after TACE therapy and then the potential mechanism and targets will be revealed. This research project investigates innovatively the mechanism of HIF-1α/miR-210 pathway from hypoxia and tumor vascular normalization, and it has an important theoretical and practical significance in depth understanding for the prevention and treatment of angiogenesis of hepatocellular carcinoma after TACE therapy and promoting the development of innovative drugs for targeting interventions for angiogenesis in hepatocellular carcinoma.

肝癌介入治疗后肿瘤血管生成严重影响预后,HIF-1α/miR-210参与肝癌血管生成的调节。本项目前期研究发现肝癌经抑制血管生成干预后存在肿瘤血管正常化期。在此基础上，针对肿瘤血管正常化期间促血管生成因子和血管生成表达相关因子表达下降，而抑制血管生成因子表达升高，提出“HIF-1α/miR-210极可能参与了血管正常化的信号通路调节，可能通过抑制促血管生成因子表达和或促进抑制血管生成因子表达来完成”科学假说，采用乏氧肝癌细胞和肝癌裸鼠移植瘤，ACI大鼠模型，以共聚焦显微镜及电镜观察肿瘤血管形态，借助siRNA等技术干预HIF-1α/miR-210，研究HIF-1α/miR-210调控肝癌血管正常化，揭示其作用机制和靶标。本项目创新地从乏氧和肿瘤血管正常化角度研究HIF-1α/miR-210的作用机制，对于阐明介入治疗后肝癌新生血管形成机制和推动靶向干预肝癌血管生成的药物研发有重要意义。

肝癌介入治疗后肿瘤血管生成严重影响预后。其可能病理机制在于肿瘤血管生成是肿瘤发展和转移的必要基础，但其调控机制尚未完全清楚。相关基因的异常表达是肝癌恶性生物学行为的基础，因此研究调控该类基因的分子是目前研究的重点。miRNA作为mRNA的上游调控因子，调控着肿瘤发生和转移相关基因的表达，miR-210即是该类重要的调控分子，且miR-210与肝癌介入治疗过程中缺氧(HIF-1ɑ)密切相关。因此，本项目从研究的源头（病人临床标本收集）到功能调节机制（信号通路），系统的探讨了HIF-1ɑ/miR-210在肝癌血管生成中的作用及相关的分子调节机制。本课题组收集了48例接受肝癌介入治疗肝癌患者血液样本，采用外泌体提取分析及体外细胞试验等方法，确认肝癌患者来源外泌体中高表达HIF-1ɑ/miR-210，并通过激活AKT通路来促进肝癌血管内皮细胞的增生及管腔形成。此外，课题组还收集了90例肝癌手术标本及对应癌旁标本，通过基因芯片筛选及鉴定，确认miR-210为肝癌促癌基因，与临床血管浸润及不良预后有关，并确认了miR-210下游靶分子CPEB2，分析及确立了HIF-1ɑ/miR-210/CPEB2环路参与调节EMT进程影响肝癌的侵袭转移。所有体外生物学实验结果均通过动物体内实验进一步验证。与此同时，我们分析了接受介入治疗患者血清miR-210及炎症因子的表达水平及术后疗效分析，发现miR-210联合炎症因子MCP-1可以有效预测介入疗效，为今后介入治疗、筛选肝癌介入治疗的适宜人群提供了充实的临床证据。本项目顺利完成miR-210在肝癌血管生成作用及其参与的复发转移机制研究的目标，同时产生原创性论文及手稿3篇，培养硕士研究生2名，以发言和壁报参加国内国际学术交流多次。